1,3-/1,4-thiomorpholino-terminated non-peptidyl α-succinamidoacyl aminodiols as anti-hypertensive agents

ABSTRACT

Non-peptidyl compounds characterized generally as α-succinamidoacyl aminodiols having a 1,3-thiomorpholino-type group or a 1,4-thiomorpholino-type group at the N-terminus are useful as renin inhibitors for the treatment of hypertension.

RELATED APPLICATION

This is a divisional of U.S. application Ser. No. 07/732,880, filed Jul.19, 1991, which is a continuation of Ser. No. 07/103,623, file Oct. 1,1987, now abandoned.

FIELD OF THE INVENTION

Renin-inhibiting compounds are known for control of hypertension. Ofparticular interest herein are non-peptidyl compounds useful as renininhibiting agents.

BACKGROUND OF THE INVENTION

Renin is a proteolytic enzyme produced and secreted into the bloodstreamby the juxtaglomerular cells of the kidney. In the bloodstream, renincleaves a peptide bond in the serum protein angiotensinogen to produce adecapeptide known as angiotensin I. A second enzyme known as angiotensinconverting enzyme, cleaves angiotensin I to produce the octapeptideknown as angiotensin II. Angiotensin II is a potent pressor agentresponsible for vasoconstriction and elevation of cardiovascularpressure. Attempts have been made to control hypertension by blockingthe action of renin or by blocking the formation of angiotensin II inthe body with inhibitors of angiotensin I converting enzyme.

Classes of compounds published as inhibitors of the action of renin onangiotensinogen include renin antibodies, pepstatin and its analogs,phospholipids, angiotensinogen analogs, pro-renin related analogs andpeptide aldehydes.

A peptide isolated from actinomyces has been reported as an inhibitor ofaspartyl proteases such as pepsin, cathepsin D and renin [Umezawa et al,in J. Antibiot. (Tokyo), 23: 259-262 (1970)]. This peptide, known aspepstatin, was found to reduce blood pressure in vivo after theinjection of hog renin into nephrectomized rats [Gross et al, Science,175, 656 (1971)]. Pepstatin has the disadvantages of low solubility andof inhibiting acid proteases in addition to renin. Modified pepstatinshave been synthesized in an attempt to increase the specificity forhuman renin over other physiologically important enzymes. While somedegree of specificity has been achieved, this approach has led to ratherhigh molecular weight hepta- and octapeptides [Boger, et al, Nature,303, 81 (1983)]; high molecular weight peptides are generally consideredundesirable as drugs because gastrointestinal absorption is impaired andplasma stability is compromised.

Short peptide aldehydes have been reported as renin inhibitors [Kokubuet al, Biochem. Biophys. Res. Common., 118, 929 (1984); Castro et al,FEBS Lett., 167, 273 (1984)]. Such compounds have a reactive C-terminalaldehyde group and would likely be unstable in vivo.

Other peptidyl compounds have been described as renin inhibitors. EPAppl. #128,762, published Dec. 18, 1984, describes dipeptide andtripeptide glycol-containing compounds as renin inhibitors [also seeHanson et al, Biochem. Biophys. Res, Comm., 132:155-161 (1985),146:959-963 (1987)]. EP Appl. #181,110, published May 14, 1986,describes dipeptide histidine derivatives as renin inhibitors. EP Appl.#189,203, published Jul. 30, 1986, describes peptidylaminodiols as renininhibitors. EP Appl. #200,406, published Dec. 10, 1986, describes alkylnaphthylmethyl propionyl-histidyl aminohydroxy alkanoates as renininhibitors.

For other articles describing previous efforts to devise renininhibitors, see Marshall, Federation Proc., 35: 2494-2501 (1976); Burtonet al, Proc. Natl. Acad. Sci. USA, 77: 5476-5479 (1980); Suketa et al,Biochemistry, 14: 3188 (1975;) Swales, Pharmac. Ther., 7: 173-201(1979); Kokubu et al, Nature, 217: 456-457 (1986); Matsushita et al, J.Antibiotics, 28: 1016-1018 (1975); Lazar et al, Biochem. Pharma,,23:2776-2778 (1974); Miller et al., Biochem. Pharma., 21: 2941-2944(1972); Haber, Clinical Science, 59: 7s-19s (1980); Rich et al, J. Org.Chem., 43: 3624 (1978); J, Med. Chem., 23: 27 (1980); especially Haber,Clin. and Exper. Hyper., A5(7&8), 1193 (1983); and European PatentApplications 172346A and 172347A published Feb. 26, 1986.

DESCRIPTION OF THE INVENTION

Non-peptidyl α-succinamidoacyl aminodiols compounds having utility asrenin inhibitors for treatment of hypertension in mammals constitute afamily of compounds of general Formula I: ##STR1## wherein X is selectedfrom ##STR2## wherein Y and Q are selected from CH₂, ##STR3## O, S, SO,SO₂ and NR₁₀, wherein R₉ is H or lower alkyl, R₁₀ is selected from H,phenyl and and wherein R₁₁ is H or lower alkyl; wherein each of m and nis independently an integer from 1 through 4; wherein each of r, t, uand v is independently an integer from zero through two; wherein p is aninteger from 1 through 3; wherein each of a through d is independentlyan integer from zero through 3; wherein T is selected from one or moregroups selected from H, linear or branched lower alkyl, alkoxy, oxo,halo, haloalkyl, lower alkenyl, lower alkynyl and cyano; wherein R₁ isselected from H, linear or branched lower alkyl, haloalkyl,alkylcycloalkyl, alkylcycloalkenyl and alkoxycarbonyl; wherein R₂ isselected from linear or branched lower alkyl and benzyl; wherein R₃ isselected from lower alkyl, acylaminoalkyl, benzyl, naphthylmethyl, aryland benzyl substituted at the phenyl portion by halo or lower alkyl orby both; wherein each of R₄ and R₅ is independently selected from H orlower alkyl; wherein R₆ is selected from substituted or unsubstitutedcycloalkyl, phenyl, cycloalkylalkyl and phenylalkyl, any one of whichmay be substituted with one or more groups selected from alkyl, alkoxy,halo, haloalkyl, lower alkenyl, lower alkynyl and cyano; and whereineach of R₇ and R₈ is independently selected from the groups H, loweralkyl, cycloalkyl, phenyl, benzyl, naphthyl and naphthylmethyl, any oneof which groups having a substitutable position may be optionallysubstituted with or more of lower alkyl, alkoxy, alkenyl, alkynyl, halo,haloalkyl, cyano and phenyl, with the proviso that at least one of R₇and R₈ is an aryl group.

A preferred group of compounds within Formula I are those compoundshaving the specific stereochemical configuration shown in Formula II:##STR4##

Preferred compounds within Formula II are those compounds of Formula Iwherein Y and Q are selected from O, CH₂, S, NR₁₀ wherein R₁₀ is H or##STR5## wherein R₁₁ is H or lower alkyl; wherein each of m and n isindependently an integer from 1 through 3; wherein each of r, t, u and vis independently zero or one; wherein p is one or two; wherein each of athrough d is independently an integer from zero through 2; wherein T isselected from one or more groups selected from H, lower alkyl, alkoxy,oxo and halo; wherein R₁ is selected from H, lower alkyl,alkylcycloalkyl and alkoxycarbonyl; wherein R₂ is selected from loweralkyl and benzyl; wherein R₃ is selected from lower alkyl,acylaminoalkyl, benzyl, napthylmethyl, aryl and benzyl substituted atthe phenyl portion by halo or lower alkyl or by both; wherein R₄ isselected from H and lower alkyl; wherein R₅ is H lower alkyl; wherein R₆is selected from cyclohexylmethyl and benzyl; wherein each of R₇ and R₈is independently selected from H, phenyl, naphthyl and phenylsubstituted with one or more lower alkyl, alkoxy, alkenyl, halo, cyanoand phenyl, with the proviso that at least one of R₇ and R₈ is phenyl.

Within the aforementioned preferred group of compounds, there are foursub-groups of preferred compounds. The first sub-group consists of thosecompounds of Formula II wherein X is ##STR6## wherein Y is O, CH₂, or S;m is 2; n is 2; T is one or more of H or lower alkyl; R₁ is H or methyl;or lower alkyl; R₂ is lower alkyl; R₃ is benzyl; R₄ is H; R₅ is H; andR₆ is cyclohexylmethyl. Of this first sub-group the most preferred arethose compounds wherein Y is O; m is 2; n is 2; T is one or more of H ormethyl; R₁ is selected from H, methyl, ethyl and isobutyl; R₂ isisobutyl; R₃ is selected from benzyl and napthylmethyl; R₄ is H ormethyl; R₅ is H or methyl; and R₆ is cyclohexylmethyl. Radicals whichexemplify the X substituent of Formula III are as follows: ##STR7##

A second sub-group of preferred compounds consists of those withinFormula I wherein X is ##STR8## wherein Y is selected from O, S, CH₂, SOand SO₂ ; wherein each of r, t, u and v is independently zero or one; pis 1 or 2; T is one or more of H, lower alkyl and alkoxy; R₁ is loweralkyl; R₂ is lower alkyl; R₃ is benzyl; R₄ is H or methyl; R₅ is H ormethyl; and R₆ is cyclohexylmethyl. The substituent of Formula IV may besubstituted at any substitutable position within the bicyclic structureof Formula IV. Radicals which exemplify the X substituent of Formula IVare as follows: ##STR9##

A third sub-group of preferred compounds consists of those compoundswherein X is ##STR10## wherein Q is O, CH₂, or S; a is zero; b is 1; cis 1; d is zero; each T is independently one or more of H or loweralkyl; R₁ is H or lower alkyl; R₂ is lower alkyl; R₃ is benzyl; R₄ is Hor methyl; R₅ is H or methyl; and R₆ is cyclohexylmethyl.

Within this third sub-group is a set of more preferred compounds ofFormula II wherein Q is O, R₁ is selected from H, methyl, ethyl andisobutyl, and R₂ is isobutyl. Especially preferred is a compound whereinR₁ is isobutyl and R₆ is cyclohexylmethyl. Another set of more preferredcompounds within this third sub-group are those wherein Q is S, R₁ isselected from H, methyl and isobutyl, and R₂ is isobutyl. Especiallypreferred is a compound wherein R₁ is isobutyl and R₆ iscyclohexylmethyl. Radicals which exemplify the X substituent of FormulaV are as follows: ##STR11##

Within any of these radicals exemplifying Formulae III, IV and V, thesubstituent R represents a linear or branched alkyl group of one toabout ten carbon atoms, or preferably, one to about five carbon atoms.

A fourth sub-group of preferred compounds consists of those compounds ofFormula II wherein X is ##STR12## wherein each of R₇ and R₈ isindependently selected from the groups H, lower alkyl, cycloalkyl,phenyl, benzyl, naphthyl, and naphthylmethyl, any one of which groupshaving a substitutable position may be optionally substituted with ormore of lower alkyl, alkoxy, alkenyl, alkynyl, halo, haloalkyl, cyanoand phenyl, with the proviso that at least one of R₇ and R₈ is an arylgroup.

Of this fourth subgroup, more preferred compounds are those wherein R₁is isobutyl, R₂ is isobutyl, R₃ is benzyl, R₄ is H or methyl, R₅ is H ormethyl, R₆ is cyclohexymethyl, and each of R₇ and R₈ is independentlyselected from, H, lower alkyl and phenyl, with at least one of R₇ and R₈being phenyl. An especially preferred compound is wherein R₇ is H and R₈is phenyl.

Unless otherwise described, the chemical groups recited herein shallhave meanings as follows: "Lower alkyl" means alkyl radicals containingone to about 10 carbon atoms in a linear or branched configuration,examples of which include methyl, ethyl, n-propyl, isopropyl, n-butyl,sec-butyl, iso-butyl, tert-butyl, n-pentyl, neopentyl, n-hexyl,1-methylhexyl, n-heptyl, 2-ethylheptyl, n-octyl, 3-propyloctyl, n-nonyl,4-butylnonyl, n-decyl and the like. "Lower alkenyl" and "lower alkynyl"mean, respectively, hydrocarbon radicals of two to about ten carbonatoms containing at least one carbon-carbon double bond and at least onecarbon-carbon triple bond, respectively, attached to alkyl radicals ofthe type embraced by the term "lower alkyl" examples of which are2-butenyl and 3-pentenyl. "Haloalkyl" means alkyl radicals substitutedat one or more substitutable positions with one or more halo groups.Preferred haloalkyl group are those provided by lower alkyl radicalssubstituted at least at one position with one, two or three halo groupssuch as fluoro or chloro, a specific example of which istrifluoromethyl. "Alkylcycloalkyl" means a cyclized alkyl having fromfour to about nine ring carbon atoms, any one or more of thesubstitutable ring carbons being substituted with an alkyl group,preferably a lower alkyl group. "Alkylcycloalkenyl" means a cyclizedhydrocarbon radical containing four to about nine ring carbon atomscontaining at least one carbon-carbon double bond, but less than thenumber of double bonds required to form a fully unsaturated ring system,any one or more of the substitutable ring carbon atoms being substitutedwith an alkyl group, preferably a lower alkyl group. "Alkoxycarbonyl"means an oxycarbonyl radical having an alkyl, preferably lower alkyl,group attached to the oxygen atom. "Aryl" means an aromatic hydrocarbonradical provided by a homocyclic ring system, such as phenyl andnaphthyl. "Acyl" means a carbonyl radical attached to a hydrocarbonmoiety, typically an alkyl or lower alkyl group.

Based upon the foregoing, the meanings of the following terms should bereadily discernible, namely, "acylaminoalkyl", "cycloalkyl","cycloalkylalkyl", "phenylalkyl" and "alkoxy".

In the cyclic structures of Formulae III, IV and V where the substituentT is shown, it is intended that the T substituent represents one or moresubstituents which may be attached at any substitutable position on anyof the described cyclic structures.

Compounds of Formula I may have two or more carbon atoms providingasymmetric sites which are important for conferring activity. Preferredcompounds have three asymmetric carbons which tend to confer improvedactivity. Such compounds whether in their pure isomer form or ascomponents of racemic mixtures are embraced in the Formula I and IIcompounds of the invention. Many of the more active renin inhibitors areprovided by compounds having a specific stereochemical configuration.Within Formula I, reading from the N terminus to the C terminus(terminating with the diol moiety), the preferred configurations for theasymmetric carbons are as follows: R,S,S,R,S.

Compounds of Formula I have been found to inhibit the production ofangiotensin II in mammals. Angiotensin II is a potent vasoconstrictorand participates in the formation of aldosterone which regulates sodiumand water balance in mammals. Thus, compounds of Formula I aretherapeutically useful in methods for treating hypertension byadministering to a hypertensive patient a therapeutically-effectiveamount of a compound of Formula I.

These compounds can be formulated into pharmaceutically-acceptabledosage forms by any of a number of well-known carriers or diluents. Thecompounds can be formulated using pharmacologically-acceptable acidaddition salts and can be used in a suitable hydrated form. Theformulated compounds can be administered in oral dosage forms such astablets, capsules, pills, powders, or granules. The compounds can alsobe administered intramuscularly, using forms known to the pharmaceuticalart. In general, the preferred form of administration is oral. Atherapeutically-effective but non-toxic quantity of the compound isemployed in treatment of high blood pressure in mammals. Thedosageregimen for preventing or treating hypertension with the compoundsof Formula I is selected upon consideration of a variety of factors,including the type, age, weight, sex, and medical condition of thepatient, the severity of the hypertension, the route of administration,and the particular compound employed. Dosages of the compounds areordinarily in the range from about 0.5 to about 100 mg/kg (activecompound-to-body weight), and preferably from about 1.0 to about 20mg/kg given orally or by injection.

Compounds of Formula I are also useful as diagnostic agents foridentification of hypertension due to renin excess.

Compounds of Formula I can be administered as prodrugs. Preferably,esterification of one or more of the hydroxyl groups of the compounds ofFormula I is accomplished with amino acids to make aminoesters,succinates to make succinic acid esters, or phosphates to makephosphoric acid esters. Aminoesters of the Formula I compounds are morepreferred.

Procedures for preparation of compounds of Formula I are set forth inthe schemes and descriptions under Generic Synthesis I and GenericSynthesis II, taken with the specific procedures described in Examples1-9 which follow thereafter. The substituents X and R₁ through R₆ are asdescribed above for the Formula I substituents. ##STR13## GenericSynthetic Description I

An allylic acetate, appropriately substituted and suitably protected, asshown in the scheme is used as starting material. This substance, andlike substances, are ozonized under standard conditions (lowtemperature, as methanol-methylene chloride solutions) and the reductionof the ozonide to an aldehyde is effected with dimethyl sulfide. Onceobtained, this type of aldehyde is treated with organometallic reagentscapable of delivering an alkyl group to the aldehyde to produce diols ofthe type shown. These diols may then be converted, using standardpeptide coupling methodology to renin inhibitors as shown via couplingto the general acid shown in the scheme. The initially obtained diol mayalso be hydrogenated to the saturated cyclohexane diol and again,coupled to in a similar manner to acids of the general description givenin the scheme.

Generic Synthetic Description II

Diols are obtained as before, but using this methods, stepwise couplingis carried out, using standard methodology developed for peptidesynthesis, to obtain the renin inhibitors depicted in the scheme.

The following Synthetic Scheme is a more specific description of thepreceeding Generic Synthesis Outlines I and II. This Synthetic Schemeoutlines preparations of the specific compounds of Examples 1-9, whichfollow.

The following examples are provided to illustrate the present inventionand are not intended to limit the scope thereof. Those skilled in theart will readily understand that known variations of the conditions andprocesses of the following preparative procedures can be used to preparethese compounds. All temperatures expressed are in degrees Centigrade.Within the foregoing synthetic description and examples which follow,abbreviations have meanings as indicated below:

BOC=butyloxycarbonyl

i-Bu=isobutyl

Leu=leucine

Ac=acyl

Me=methyl

TFA=trifluoroacetic acid

THF=tetrahydrofuran

EXAMPLE 1

(3S, 4S)-N-[(tert-Butyloxy)carbonyl]-4-amino-3-acetoxy-5-phenylpentene

The preparation of the above intermediate was carried out using theprocedure described in Hanson, et al., (1985 ) J. Org. Chem. 50,5399.

EXAMPLE 2

(2R,3S)-N-[(tert-Butyloxy)carbonyl]-3-amino-2-acetoxy-4-phenylbutanal

The preparation of the above intermediate was carried out as describedin Hanson, et al. above. Ozone/oxygen was bubbled at -70° into asolution of 2.55 g (8.0 mmol) of the allylic acetate of Example 1 in 100mL of methylene chloride until a deep blue color persisted. Oxygen wasintroduced until the blue color completely faded, then 3.0 mL of Me₂ Swas added and the solution was allowed to warm to 0°-5° and standovernight. The solvent was removed at 0° under vacuum yielding the titlecompound as a thick yellow oil which was used in the following stepwithout purification.

EXAMPLE 3

(2S, 3R,4S)-N-[(tert-Butyloxy)carbonyl]-2-amino-1-phenyl-3,4-dihydroxy-6-methylheptane

The oil prepared in Example 2 was dissolved under nitrogen in 100 mL ofdry THF and cooled to -70°. To this solution was added 13 mL (26 mmol)of a 2. OM solution of isobutylmagnesium chloride in ether and thestirred mixture was allowed to warm to room temperature and stir for 2hrs. After decomposition with MeOH/H₂ O the mixture was diluted withether, washed with saturated NH₄ Cl solution twice, then dried and thesolvents stripped off under vacuum. The residue was allowed to standovernight in 80% MeOH--H₂ O containing excess ammonium hydroxide. TheMeOH was stripped off and the mixture was extracted with ether. Theseextracts were combined, washed with water, dilute KHSO₄, then dried andevaporated to give 2.36 g of a yellow glass which crystallized from 50mL of pentane on standing overnight. The yellow-white powder obtainedwas recrystallized from ether-hexane and furnished the title compound(0.41 g) as white, hairy needles, mp 134°-136° , Rf (ether): singlespot, 0.6. By chromatography of the mother liquors and crystallizationof the appropriate fractions, an additional 0.22 g of product, mp138°-139° was obtained.

Anal: Calcd. for C₁₉ H₃₁ NO₄ (337.45): C, 67.62; H, 9.26; N, 4.15.Found: C, 67.51; H, 9.43; N, 4.24.

EXAMPLE 4

(2S,3R,4S)-N-[(tert-Butyloxy)carbonyl]-2-amino-1-cyclohexyl-3,4-dihydroxy-6-methylheptane

The diol of Example 3, 0.27 g, was reduced in MeOH with 60 psi H₂ at 60°in 3 hrs using 5% Rh/C catalyst. After filtering, the solvent wasstripped off and the 0.27 g of white crystals were recrystallized fromCH₂ Cl₂ -hexane to furnish tiny needles of the title compound, 0.19 g,mp 126°-128° ; further recrystallization gave mp 128.5°-129.5° Rf(ether): single spot, 0.8.

Anal: calcd. for C₁₉ H₃₇ NO₄ (343.50): C, 66.43; H, 10.86, N, 4.08.Found: C, 66.43; H, 11.01; N, 4,03.

EXAMPLE 5

(2S,3R)-Boc-L-Leucinamide of 2-amino-1-cyclohexyl-3,4-dihydroxybutane

The procedure of Example 4 was employed using(2S,3R)-N-Boc-2-amino-1-phenyl-3,4-dihydroxybutane, prepared by themethod of Hanson, et al. (1985) J. Org. Chem. 50, 5399-540, to give thecrystalline 1-cyclohexyl derivative:

Anal. calcd for C₁₅ H₂₉ NO₄ : C,62.69; H,10.17: N,4.87. Found: C62.69;H,10.51; N,4.91. This compound was treated with TFA and coupled toBoc-L-leucine using the coupling procedure in Example 9 to give thetitle compound.

EXAMPLE 6

D,L-Monomethyl-2-(1-naphthylmethyl)succinate

To a solution of diisopropylamine (36.6 g, 362 mmol) in dry THF (200ml)cooled to -78° for 30 minutes. To this solution was added monomethylsuccinate (23.8 g, 180 mmol) in THF (200 mL) over a 30 minute period,and the reaction mixture was allowed to stir at -78° for 1 hour. To thismixture, 1-(bromomethyl)naphthylene (40 g, 191 mmol) in THF (220 mL) wasadded over a 30 minute period, and the solution was stirred for 1 hourat -78° then room temperature for 18 hours. The moisture was dilutedwith ether and extracted with water. The aqueous extracts were combinedand acidified to pH 1 with 10% aqueous HCl, extracted with either andthe organic extracts dried over sodium sulfate, filtered, and thesolvent evaporated. The yellow solid residue was dissolved in hottoluene and hexane was added until the solution become cloudy. Thesolution was cooled, filtered and the title compound was isolated as apale yellow solid (21 g). Anal. calcd for C₁₆ H₁₆ O₄ : C,70.58; H,5.92.Found: C,69.03; H,5.99.

EXAMPLE 7

D,L-Methyl 3-(N-morpholinocarbonyl)-2-(1-naphthylmethyl)-propionate

The title compound of Example 6 (6 g, 22 mmol was dissolved in methylenechloride (50 mL) and the solution cooled to 0°. Thionyl chloride (10.5g, 88 mmol) was added over a 2 minute period. The solution was thenallowed to warm to room temperature and was stirred for 2 hours. Solventand excess thionyl chloride were removed under vacuum and the residuewas dissolved in methylene chloride. To this solution was addedmorpholine (5.75 g, 66 mmol) in methylene chloride (50 mL) over a 15minute period. The mixture was stirred at room temperature for 18 hours,washed with 10% aqueous HCl followed by saturated aqueous potassiumhydrogen carbonate, dried over sodium sulfate and chromatographed onsilica gel eluting with 30/70 ethyl acetate-methylene chloride to obtainthe title compound (4.48 g):

Anal. calcd: C₂₀ H₂₃ O₄ : C,71.20; H,5.68; N,4.15. Found: C,69.12:H,7.03; N,3.79.

EXAMPLE 8

D,L-3-(N-Morpholinocarbonyl)-2-(1-naphthylmethyl)propionic acid

The title compound of Example 7 (4.38 g, 8 mmol) was dissolved inmethanol (25 mL) and treated with aqueous sodium hydroxide (2N, 12.87mL). The resulting solution was stirred for 6 hours at room temperature,then the solution was reduced to a small volume in vacuo and the residuewas taken up in water. Concentrated aqueous HCl was added dropwise untilpH 1 was reached, then the mixture was extracted with ethyl acetate. Theorganic extracts were dried over sodium sulfate and the solventevaporated to obtain the title compound as a solid (4.15 g):

Anal. calcd for C₁₉ H₂₁ O₄ : C,69.71: H,6.47; N,4.28. Found C,68.47;H,6.74; N,3.96.

EXAMPLE 9

3-(N-morpholinocarbonyl)-2-(R,S)-(1-naphthylmethyl)propionyl-L-leucinamideof (2S ,3R)-2-amino-1-cyclohexyl-3,4-dihydroxybutane

The title compound of Example 5 was treated with trifluoroacetic acid toremove the Boc group and this salt (96 mg) was dissolved in methylenechloride and treated with N-methylpiperidine (23 mg) to form Solution A.The title compound of Example 8 (109 mg) was dissolved in methylenechloride and treated with N-methylpiperidine (33 mg), then cooled to-10. To this solution was added isobutylchloroformate (43 mg) and after4 minutes, Solution A was added. This mixture was allowed to stir at-10° for 4 hours and then the solvent was evaporated to give an oilyresidue. This was taken up in methanol and 1N potassium hydroxide wasadded; after 5 minutes, 0.5M citric acid was added and the mixtureextracted with ethyl acetate. The organic layer was washed with 5%aqueous potassium carbonate, brine, and dried over sodium sulfate togive the title compound as a white foam (97 mg): 400 MH_(z) ¹ H NMR(CDCl₃): consistent with proposed structure:

Anal. calcd for C₃₅ H₅₁ N₃ O₆ +0.75 H₂ O: C,67.44; H,8.48; N,6.74.Found: C,67.37; H,8.25; N,6.42.

Biological Evaluation:

Compounds of Formula I were evaluated as inhibitors of human renin in anin vitro assay, as follows. This human renin inhibition test has beenpreviously described in detail [Papaioannou, et al., Clinical andExperimental Hypertension, A7(9), 1243-1257 (1985)]. Human renin wasobtained from the National Institute for Biological Standards, London.In a total volume of 0.25 mL 100 mM Tris-acetate buffer at pH 7.4,25×10-6 Goldblatt units of renin, 0.05 mL of plasma from humanvolunteers taking oral contraceptives, 6.0 mM sodium EDTA, 2.4 mMphenylmethyl sulfonyl fluoride, 1.5 mM 8-hydroxyquinoline, 0.4 mg/mLBSA, and 0.024 mg/mL neomycin sulfate were incubated for two hours at37° C. in the presence or absence of renin inhibitors. The producedangiotensin I was determined by radioimmunoassay (New England Nuclearkit). Compounds to be assayed were solubilized in either ethyl alcoholor DMSO and diluted with 100 mM Tris-acetate buffer at pH 7.4 to theappropriate concentration. The final concentration of organic solvent inthe reaction mixture was less than 1%. Control incubations at 37° C.were used to correct for effects of organic solvent on renin activity.

The hog renin inhibition assay was performed in a manner similar to thehuman renin assay, with the following modifications. Hog renin waspurchased from Sigma Chemical Co. and the synthetic tetradecapeptidesubstrate was obtained from Peninsula Labs Inc. In a final volume of0.25 mL 100 mM Tris-acetate buffer at pH 7.4, 0.125 m units hog renin,20 micromolar tetradecapeptide, 6 mM disodium EDTA, 3.2 mM phenylmethylsulfonyl fluoride, 3 mM 8-hydroxyquinoline, 1.2 mg/mL BSA and 0.024mg/mL neomycin sulfate were incubated for one hour at 37° C. in thepresence or absence of renin inhibitors. The amount of angiotensin Iproduced was determined as for the human renin inhibition assay.Biological Results:

                  TABLE I                                                         ______________________________________                                        Compound    Human Renin IC.sub.50                                                                       Hog Renin IC.sub.50                                 ______________________________________                                        Example 9   7.4 × 10.sup.-7 M                                                                     2.8 × 10.sup.-6 M                             ______________________________________                                    

Although this invention has been described with respect to specificembodiments, the details of these embodiments are not to be construed aslimitations. Various equivalents, changes and modifications may be madewithout departing from the spirit and scope of this invention, and it isunderstood that such equivalent embodiments are part of this invention.

what is claimed is:
 1. A compound of the formula: ##STR14## wherein x isselected from ##STR15## wherein T is selected from one or more groupsselected from linear or branched lower alkyl, lower alkoxy, oxo, halo,haloloweralkyl, lower alkenyl, lower alkynyl and cyano; wherein R₁ isselected from linear or branched lower alkyl, haloloweralkyl, loweralkylcycloalkyl, lower alkylcycloalkenyl and lower alkoxycarbonyl;wherein R₂ is selected from linear or branched lower alkyl and benzyl;wherein R₃ is selected from lower alkyl, lower alkylcarbonylaminoalkyl,benzyl, naphthylmethyl, phenyl, naphthyl and benzyl substituted at thephenyl portion by halo or lower alkyl or by both; wherein each of R₄ andR₅ is independently selected from H or lower alkyl; and wherein R₆ isselected from substituted or unsubstituted cycloalkyl, phenyl,cycloalkylalkyl and phenylalkyl, any one of which may be substitutedwith one or more groups selected from lower alkyl, lower alkoxy, halo,haloloweralkyl, lower alkenyl, lower alkynyl and cyano.
 2. Compound ofclaim 1 of the Formula: ##STR16## wherein X and R₁ through R₆ are asdefined in claim
 1. 3. Compound of claim 2 wherein T is selected fromone or more groups selected from lower alkyl, lower alkoxy, oxo andhalo; wherein R₁ is selected from lower alkyl, lower alkylcycloalkyl andlower alkoxycarbonyl; wherein R₂ is selected from lower alkyl andbenzyl; wherein R₃ is selected from lower alkyl, loweralkylcarbonylaminoalkyl, benzyl, naphthylmethyl, phenyl, naphthyl andbenzyl substituted at the phenyl portion by halo or lower alkyl or byboth; wherein R₄ is selected from H and lower alkyl; wherein R₅ is H orlower alkyl; and wherein R₆ is selected from cyclohexylmethyl andbenzyl.
 4. Compound of claim 3 wherein T is one or more of lower alkyland lower alkoxy; wherein R₁ is H or lower alkyl; wherein R₂ is loweralkyl; wherein R₃ is benzyl; wherein R₄ is H or methyl;: wherein R₅ is Hor methyl; and wherein R₆ is cyclohexylmethyl.
 5. A pharmaceuticalcomposition comprising a therapeutically-effective amount of arenin-inhibiting compound and a pharmaceutically-acceptable carrier ordiluent, said renin-inhibiting compound selected from a family ofcompounds of the formula: ##STR17## wherein X is selected from ##STR18##wherein T is selected from one or more groups selected from linear orbranched lower alkyl, lower alkoxy, oxo, halo, haloloweralkyl, loweralkenyl, lower alkynyl and cyano; wherein R₁ is selected from linear orbranched lower alkyl, haloloweralkyl, lower alkylcycloalkyl, loweralkylcycloalkenyl and lower alkoxycarbonyl; wherein R₂ is selected fromlinear or branched lower alkyl and benzyl; wherein R₃ is selected fromlower alkyl, lower alkylcarbonylaminoalkyl, benzyl, naphthylmethyl,phenyl, naphthyl and benzyl substituted at the phenyl portion by halo orlower alkyl or by both; wherein each of R₄ and R₅ is independentlyselected from H or lower alkyl; and wherein R₆ is selected fromsubstituted or unsubstituted cycloalkyl, phenyl, cycloalkylalkyl andphenylalkyl, any one of which may be substituted with one or more groupsselected from lower alkyl, lower alkoxy, halo, haloloweralkyl, loweralkenyl, lower alkynyl and cyano.
 6. The composition of claim 5 whereinT is selected from one or more groups selected from lower alkyl, loweralkoxy, oxo and halo; wherein R₁ is selected from lower alkyl, loweralkylcycloalkyl and lower alkoxycarbonyl; wherein R₂ is selected fromlower alkyl and benzyl; wherein R₃ is selected from lower alkyl, loweralkylcarbonylaminoalkyl, benzyl, naphthylmethyl, phenyl, naphthyl andbenzyl substituted at the phenyl portion by halo or lower alkyl or byboth; wherein R₄ is selected from H and lower alkyl; wherein R₅ is H orlower alkyl; and wherein R₆ is selected from cyclohexylmethyl andbenzyl.
 7. The composition of claim 6 wherein T is one or more of loweralkyl and lower alkoxy; wherein R₁ is H or lower alkyl; wherein R₂ islower alkyl; wherein R₃ is benzyl; wherein R₄ is H or methyl; wherein R₅is H or methyl; and wherein R₆ is cyclohexylmethyl.
 8. A therapeuticmethod for treating hypertension, said method comprising administeringto a hypertensive patient a therapeutically-effective amount of acompound of the Formula: ##STR19## wherein X is selected from ##STR20##wherein T is selected from one or more groups selected from linear orbranched lower alkyl, lower alkoxy, oxo, halo, haloloweralkyl, loweralkenyl, lower alkynyl and cyano; wherein R₁ is selected from linear orbranched lower alkyl, haloloweralkyl, lower alkylcycloalkyl, loweralkylcycloalkenyl and lower alkoxycarbonyl; wherein R₂ is selected fromlinear or branched lower alkyl and benzyl; wherein R₃ is selected fromlower alkyl, lower alkylcarbonylaminoalkyl, benzyl, naphthylmethyl,phenyl, naphthyl and benzyl substituted at the phenyl portion by halo orlower alkyl or by both; wherein each of R₄ and R₅ is independentlyselected from H or lower alkyl; and wherein R₆ is selected fromsubstituted or unsubstituted cycloalkyl, phenyl, cycloalkylalkyl andphenylalkyl, any one of which may be substituted with one or more groupsselected from lower alkyl, lower alkoxy, halo, halo- loweralkyl, loweralkenyl, lower alkynyl and cyano.
 9. The method of claim 8 wherein T isselected from one or more groups selected from lower alkyl, loweralkoxy, oxo and halo; wherein R₁ is selected from lower alkyl, loweralkylcycloalkyl and lower alkoxycarbonyl; wherein R₂ is selected fromlower alkyl and benzyl; wherein R₃ is selected from lower alkyl, loweralkylcarbonylaminoalkyl, benzyl, naphthylmethyl, phenyl, naphthyl andbenzyl substituted at the phenyl portion by halo or lower alkyl or byboth; wherein R₄ is selected from H and lower alkyl; wherein R₅ is H orlower alkyl; and wherein R₆ is selected from cyclohexylmethyl andbenzyl.
 10. The method of claim 9 wherein T ms one or more of loweralkyl and lower alkoxy; wherein R₁ is H or lower alkyl; wherein R₂ islower alkyl; wherein R₃ is benzyl; wherein R₄ is H or methyl; wherein R₅is H or methyl; and wherein R₆ is cyclohexylmethyl.